Many applications in the field of analytical research and clinical testing utilize optical methods for analyzing liquid samples. Among those methods are absorbance, turbidity, fluorescence/luminescence, and optical scattering measurements. Optical laser scattering is one of the most sensitive methods, but its implementation can be very challenging, especially when analyzing biological samples in which suspended particles are relatively transparent in the medium. In this case, most of the scattering process occurs in the forward direction near the incident laser beam. To detect this low-angle, forward scattering signal, high extinction of the incident beam is required. But various optical effects (e.g., such as laser beam spatial purity, optical surface scattering, and beam distortions by the free liquid surface) often interfere with the extinction of the incident beam. For this reason, the forward scattering method is rarely applied in spite of its sensitivity.
In the case of fluorescence/luminescence detection, there is a spectral separation between the excitation light and the emitted light, which can help to facilitate the extinction of the excitation light by means of spectroscopic techniques, such as notches, bandpass optical filters, or monochromators. But in many application cases, the fluorescence signal is many orders of magnitude lower compared to the excitation light intensity and the excitation-light extinction by wavelength separation is not sufficient. For this reason, many systems collect the emitted light from a direction that is opposite of or normal to the excitation beam, such that the excitation light does not reach the detector. However, this can results in a rather complex optical layout, sometimes utilizing multiple detectors.
One particularly important application of optical measurements of liquid samples involves a microplate reader used for microbiologic assays. A microplate comprises of multiple open-top wells containing individual samples arranged in a two dimensional array (e.g., 8×12). To obtain useful information on the samples content, the microplate reader may utilize one or more types of optical measurements. Because of a two dimensional arrangement, the optical access is typically available in only the top and bottom directions of the wells. The upper free surface of the liquid sample is normally curved due to the liquid's surface tension. This curvature combined with the relatively small diameter of the wells cause a significant incident beam divergence or distortion, making its extinction very difficult and inefficient before it reaches the detector. This is one reason that forward scatter signal measurement or a fluorescence signal measurement in the input-beam direction is not easily implemented in wells of a microplate.
Accordingly, there is a need for an improved optical measurement system that allows for the detection of the forward scatter signals and/or forward fluorescence signals in the input-beam direction so as to allow for a determination of the size, quantity, and/or concentration of particles (e.g., bacteria) in the liquid.